Transparent Tissue-Visualizng Preparation

ABSTRACT

Described is a highly safe means to improve visibility of transparent tissues of the eye, i.e., the vitreous body, the lens or the cornea, in a surgical operation on them. The means comprises fine particles of a macromolecular compound, in particular, those one gram of which cannot be dissolved in less than 30 mL of water within 30 minutes at 20° C., and whose average particle size is 0.01-500 μm, and which, among others, is a biodegradable macromolecular compound and/or a water soluble macromolecular compound.

TECHNICAL FIELD

The present invention relates to a transparent tissue-visualizingpreparation which is designed to be infused into or sprinkled over atransparent tissue of the eye in order to enhance visibility oftransparent tissues of the eye in surgical operation, as well as to amethod to visualize and enhance visibility of a transparent tissue ofthe eye by application of the preparation.

BACKGROUND ART

To receive light from a surrounding environment and project it on thephotoreceptor cells, many portions of the eyeball are made oftransparent tissues, i.e., the cornea, lens and vitreous body. Thevitreous body, which adjoins the retina, provides scaffolds toproliferating tissues formed in the retina in many of retinal diseasesincluding diabetic retinitis. As tissues that have proliferated into thevitreous body will form fibers there, which draw the retina and therebycould cause retinal detachment, a condition which, if left untreated,could eventually lead to blindness. Therefore, a surgical operation isoften performed to completely remove such a vitreous body that has cometo contain proliferating tissues.

In vitrectomy, it is required to remove, thoroughly as far as possible,not only proliferating tissues that have adhered to the retina but alsothe vitreous body, which provides scaffolds for the proliferation ofsuch tissues. The operation is carried out while infusing into theeyeball an intraocular irrigating solution for surgical use. However,the vitreous body is a transparent tissue, whose refractive indexdiffers very little from that of available intraocular irrigatingsolutions. Accordingly, when no countermeasure is taken, the transparenttissue lacks visibility in the field seen through an operationmicroscope, and this makes it hard to locate the tissue, thus makingcomplete removal of it no easy task. As a means to address this problem,a steroid suspension such as a triamcinolone preparation (Kenacort-A:registered trademark) is infused during vitrectomy into a vitreouscavity, a space created by sucking and removing central part of thevitreous body, in order for letting the suspension disperse and adhereto the vitreous body and thereby visualize the vitreous body (seeNon-patent Document 1). This method improves visibility of the vitreousbody in the operative field, and thus makes the surgical operationeasier, and complete removal of the vitreous body available. However, inassociation with application of a steroid preparation to the vitreousbody, there have been reported, as their side effects, elevation of theintraocular pressure and progression of cataract (see Non-patentDocuments 2 and 3). Thus, similar side effects could be induced by asteroid suspension used during vitrectomy for the purpose of improvingvisibility.

On the other hand, there is reported a method to visualize proliferatingmembranes and the epiretinal membrane which are associated withproliferative vitreoretinopathy (PVR), by staining them with an aqueoussolution containing a dissolved dye, such as trypan blue (see PatentDocument 1). An improvement in the visibility provided by the dye thusemployed, however, is quite insufficient, since it darkens the operativefield and, besides, does not serve to emphasize the vitreous body in theoperative field.

Also in the case of cataract surgery, for example, in which the nucleusand cortex enclosed in the lens capsule are removed and then anintraocular lens is inserted, it is known that remaining cortex thatcould not be removed at and near the posterior capsule oftenproliferates and become opaque with the lapse of time after operation,thus leading to the development of postoperative cataract. Therefore,also with regard to cataract surgery, a means is needed to visualizetransparent part of the cortex and thereby make its complete removaleasier.

[Patent Document 1] WO 99/058159

[Non-patent Document 1] Sakamoto, T., et al., Graefe's Archive forClinical and Experimental Opthalmology, 240: p. 423 (2002).

[Non-patent Document 2] Challa, J. K. et al., Australian and New ZealandJournal of Opthalmology, 26: p. 277 (1998)

[Non-patent Document 3] Wingate, R. J. et al., Australian and NewZealand Journal of Opthalmology, 27: p. 431 (1999)

DISCLOSURE OF INVENTION Problem to be Solved by the Invention

Against the above-mentioned background, the objective of the presentinvention is to provide a means to improve visibility of transparenttissues of the eye, i.e., vitreous body, lens or cornea in surgicaloperations, which means enables to achieve sufficient visibility, iseasy to use and excels in safety.

Means to Solve the Problem

As a result of studies addressed to the above problem, the presentinventors found that a macromolecular compound that has nopharmacological activity and dissolves in the body fluid only graduallyand is eliminated and/or absorbed along with the turnover of the fluidin the eye, among others, particularly preferably, a biodegradablemacromolecular compound, when infused in or sprinkled over oculartransparent tissues in the form of fine particles, adheres to thesurface of those transparent tissues, on which it scatters visible lightin an operative field while it remains in the form of particles, thusserving to achieve excellent visibility without raising safety concerns.The present invention has been completed through further studies basedon the finding. Thus, the present invention provides what follows:

(1) A transparent tissue-visualizing preparation designed to be broughtinto contact with transparent tissues of the eye to enhance visibilitythereof, comprising fine particles of a macromolecular compound.

(2) The transparent tissue-visualizing preparation as defined in (1)above, wherein the fine particles are those one g of which cannotcompletely be dissolved in less than 30 mL of water within 30 minutes at20° C.

(3) The transparent tissue-visualizing preparation as defined in (1) or(2) above, wherein the average size of the particles is 0.01-500 μm.

(4) The transparent tissue-visualizing preparation as defined in one of(1) to (3) above, wherein the macromolecular compound is a biodegradablemacromolecular compound and/or a water soluble macromolecular compound.

(5) The transparent tissue-visualizing preparation as defined in (4)above, wherein the biodegradable macromolecular compound is selectedfrom fatty acid polyesters, polysaccharides and derivatives thereof.

(6) The transparent tissue-visualizing preparation as defined in (4)above, wherein the water soluble macromolecular compound is selectedfrom acrylic polymer compounds and styrene-based polymer compounds.

(7) The transparent tissue-visualizing preparation as defined in one of(1) to (3) above, wherein the macromolecular compound is selected fromthe group consisting of starch, soluble starch, pregelatinized starch,cellulose, acetylcellulose, hydroxypropylmethylcellulose, chitosan,chitin, dextran, polylactic acid, polyglycolic acid, lacticacid-glycolic acid copolymer, polyhydroxybutyric acid,polyhydroxyvaleric acid, polycaprolactone, hydroxybutyric acid-glycolicacid copolymer, lactic acid-caprolactone copolymer, polyethylenesuccinate, and polybutylene succinate, sodium polyacrylate, sodiummethacrylate and sodium polystyrenesulfonate.

(8) The transparent tissue-visualizing preparation as defined in (7)above, wherein the average molecular weight of the macromolecularcompound is 500-200000.

(9) The transparent tissue-visualizing preparation as defined in (8)above further comprising a polyvinyl-based compound and/or a polyol.

(10) The transparent tissue-visualizing preparation as defined in (9)above containing, per one part by weight of the fine particles, 0.05-10part by weight of the polyvinyl-based compound and/or 0.05-10 part byweight of the polyol.

(11) The transparent tissue-visualizing preparation as defined in (9) or(10) above, wherein the polyvinyl-based compound is polyvinylpyrrolidone and/or polyvinylalcohol.

(12) The transparent tissue-visualizing preparation as defined in (9) or(10) above, wherein the polyol is mannitol.

(13) The transparent tissue-visualizing preparation as defined in one of(1) to (12) further comprising an aqueous medium, in which the fineparticles are dispersed.

(14) The transparent tissue-visualizing preparation as defined in (13)above, wherein the content of the fine particles in the transparenttissue-visualizing preparation is 0.005-10 w/v %.

(15) The transparent tissue-visualizing preparation as defined in (13)or (14) above further containing a polyvinyl-based compound and/or apolyol.

(16) The transparent tissue-visualizing preparation as defined in (15)above containing the polyvinyl-based compound at a concentration of0.1-5 w/v % and/or the polyol at a concentration of 0.1-5 w/v %.

(17) The transparent tissue-visualizing preparation as defined in one of(1) to (16) above which is a preparation designed for injection.

(18) A transparent tissue-visualizing preparation comprising a singlemixing and discharging means which encloses, separately and in a mannerof keeping from contacting with each other, a powder compositioncomprising the fine particles as defined in one of (1) to (12) above andan aqueous medium.

(19) A method for improving visibility of a transparent tissue of theeye, which method comprises bringing a composition comprising the fineparticles as defined in one of (1) to (17) into contact with atransparent tissue of the eye.

(20) Use of the fine particles as defined in one of (2) to (8) above forthe production of a transparent tissue-visualizing preparation which isdesigned to be brought into contact with transparent tissue of the eyeto visualize it.

Effect of the Invention

In surgical operations on transparent tissues of the eye, i.e., thevitreous body, the lens and the cornea, which otherwise are hardlyvisible in the operative field, the present invention as defined aboveis brought into contact with transparent tissues and greatly enhancestheir visibility, thereby allowing easier manipulation in such surgicaloperations and, therefore, steady and easier achievement of the purposeof such operations. Since fine particles having no pharmacologicalactivity are employed, they will induce neither unnecessarypharmacological reactions nor side effects in the body. In particular,in the case where soluble and/or biodegradable fine particles areemployed, even if part of the fine particles have adhered to tissues ofthe eye and remained there after operation, they will be eliminated fromthe tissues of the eye with the lapse of time through dissolution and/ordecomposition, and then excretion or absorption, thus being less likelyto cause a problem.

BRIEF DESCRIPTION OF DRAWINGS

[FIG. 1] A schematic cross-sectional view of an example of thetransparent tissue-visualizing preparation in the form of a prefilleddouble chamber syringe.

EXPLANATION OF SIGNS

1=double chamber syringe, 2=slidable partition, 3=solid phase side,4=liquid phase side, 7=discharge flow path, 8=piston, 9=bypassing flowpath

BEST MODE FOR CARRYING OUT THE INVENTION

The transparent tissue-visualizing preparation of the present inventionis designed to enhance, by its infusion into or sprinkling over atransparent tissue of the eye, visibility of the transparent tissue inophthalmic surgery. In particular, the transparent tissue-visualizingpreparation of the present invention, when infused into (in the form ofa liquid preparation) or sprinkled over (in the form of a powder) avitreous cavity, countless numbers of its fine particles adhere to thevitreous gel, which, when irradiated with visible light, scatter thelight along the surface of the vitreous gel and thereby enhance thevisibility of the vitreous body, allowing easier visual detection of itin an operative field, e.g., through an operating microscope, duringintraocular surgeries involving removal of the vitreous body for suchdiseases as diabetic retinopathy, retinal vein occlusion, macular edema,diabetic maculopathy, macular hole, epiretinal membrane formation,rhegmatogenous retinal detachment, and so on. Also in surgicaloperations which include removal of the nucleus and cortex of the lens,the preparation, when infused into or sprinkled over the lens capsule,likewise improves the visibility of them, for a countless numbers of itsfine particles adhere to the nucleus and cortex of the lens and scatterlight along the surface of them.

In the present invention, as a “macromolecular compound”, amacromolecular compound may be employed that is pharmacologicallyinactive on ocular tissues and body of mammals, especially humans. Thereare no particular limitations as to the molecular weight of amacromolecular compound employed in the present invention, for it isenough that such a compound is solid at an ordinary temperature and canbe formed into particles. Usually, the molecular weight of it is notless than 500, preferably not less than 1000, more preferably not lessthan 1500, and particularly preferably not less than 2000. And it isusually not more than 200000, preferably not more than 150000, morepreferably not more than 100000, and particularly preferably 50000.However, those that fall outside these ranges may also be employedinsofar as they serve to achieve the objective of the present invention.

Although a variety of compounds may be used as said macromolecularcompound, it is preferred that one gram of fine particles made of agiven macromolecular compound cannot be completely dissolved in 30 mL ofwater within 30 minutes at 20° C., for such a degree of solubility issufficient for the fine particles to resist total dissolution and remainas light scattering sources during a usual course of an surgicaloperation. This degree of solubility corresponds to a solubilitycovering from “sparingly soluble”, to “slightly soluble”, “very slightlysoluble” and “practically insoluble” as defined in the JapanesePharmacopoeia, 14th edition. The Pharmacopoeia defines solubility basedon the extent to which one gram of a solute is dissolved in a solventwithin 30 minutes at 20±5° C., with stirring for 30 seconds in every 5minutes. In the present invention, this standard is substantiallyfollowed, and those fine particles are favorably employed that exhibitone of the following solubility levels when one gram of which is wellstirred in water for 30 minutes at 20° C.

(1) sparingly soluble (i.e., complete dissolution is impossible in lessthan 30 mL of water, but there exists a volume of water in the range offrom not less than 30 mL and to less than 100 mL, in which completedissolution is available.)

(2) slightly soluble (i.e., complete dissolution is impossible in lessthan 100 mL of water, but there exists a volume of water in the range offrom not less than 100 mL and to less than 1000 mL, in which completedissolution is available.)

(3) very slightly soluble (i.e., complete dissolution is impossible inless than 1000 mL of water, but there exists a volume of water in therange of from not less than 1000 mL and to less than 10000 mL, in whichcomplete dissolution is available.)

(4) practically insoluble (i.e., at least 10000 ml of water is neededfor complete dissolution).

When a macromolecular compound takes the form of a salt, preferred, butnon-limiting examples include sodium salt, potassium salt,hydrochloride, etc.

There are no clear limitations as to the average size of the fineparticles of a macromolecular compound. Considering, however, easyhandling in use and efficiency of scattering visible light, it ingeneral is preferably 0.01-500 μm, more preferably 0.1-200 μm, and stillmore preferably 1-60 μm.

Considering a possibility that the fine particles of a macromolecularcompound used partly remain in the tissues after surgical operation, itis preferred that a macromolecular compound employed is biodegradable orhas some solubility in water, because even if some of such fineparticles are left in the tissues of the eye after a surgical operation,they will be eliminated from the tissues of the eye either along withthe turnover of the liquid present in the tissues of the eye in whichthey are dissolved, after being decomposed with the lapse of time orwithout undergoing decomposition, or through absorption and furtherdecomposition.

A variety of biodegradable macromolecular compounds may be employed inthe present invention. Examples include, but not limited to, fatty acidpolyesters such as polylactic acid, polyglycolic acid, lacticacid-glycolic acid copolymer, polyhydroxybutyric acid,polyhydroxyvaleric acid, polycaprolactone, hydroxybutyric acid-glycolicacid copolymer, lactic acid-caprolactone copolymer, polyethylenesuccinate, and polybutylene succinate and like; polysaccharides andtheir derivatives such as starch and starch derivatives includingsoluble starch, pregelatinized starch and the like, cellulose andcellulose derivatives such as acetylcellulose,hydroxypropylmethylcellulose and the like, as well as chitosan, chitin,dextran and the like.

There are no clear-cut limitations as to the level of water solubilityof macromolecular compounds that may be employed in the presentinvention. For example, such a level of solubility is sufficient thatdissolution be achieved when stirred for one week with excess amount ofwater at 37° C. For, even if some portion of the fine particles are leftin the eye after a surgical operation, the amount of them will usuallybe very small and it is without problems insofar as the particlesdissolve and lose their solid form even only gradually. Examples of suchmacromolecular compounds, except for biodegradable macromolecularcompounds, include, but are not limited to, acrylic polymers andstyrene-based polymers, such as sodium polyacrylate, sodiummethacrylate, and sodium polystyrenesulfonate.

The transparent tissue-visualizing preparation of the present inventionmay contain, along with fine particles consisting of a macromolecularcompound, either one or both of polyvinyl-based compounds and polyols.Polyvinyl-based compounds and polyols act favorably to promotedispersibility of the fine particles made of a macromolecular compound.Examples of particularly preferred polyvinyl-based compounds include,but are not limited to, polyvinylpyrrolidone and polyvinylalcohol, andan example of particularly preferred polyols is mannitol. Thus, watersoluble, inactive polyvinyl-based compounds and polyols that have nopharmacological activity may be employed as desired. The amount of apolyvinyl-based compound or a polyol to be employed may be usually about0.05-10 part by weight per one part by weight of the fine particles ofthe macromolecular compound. However, their amount may fall outside thisrange, since even a lower amount of them gives some effect correspondingthereto, and higher amount of them will cause no particulardisadvantage.

The transparent tissue-visualizing preparation of the present inventionmay be in the form of a powder, or it may be in the form comprising anaqueous medium containing fine particles of a macromolecular compounddispersed in it (i.e., suspension). In the case where the preparation isin the form of a powder, it may be used by being directly sprinkled overtransparent tissues of the eye, or by being infused after mixing, priorto use, with an intraocular irrigating solution. Typically, thepreparation may be used, after suspended in a conventional manner in anintraocular irrigating-washing solution (e.g., Opeguard MA®) or OpeguardNeo Kit®, both manufactured by Senju Pharmaceutical Co., Ltd.), or in anartificial tear or so on. If the preparation is supplied in the form ofa suspension, it may be directly infused into, or applied in drops onto,transparent tissues through an injection needle.

An “aqueous medium” used for providing the transparenttissue-visualizing preparation of the present invention in the form of asuspension, is water or, as desired, a medium which is made of water andone or more additives acceptable to tissues of the eye, esp. intraoculartissues, such as isotonizers, e.g., salts and saccharides, bufferingpreparations, and the like. Examples of it include injectable distilledwater, buffered physiological saline, commercially available intraocularirrigating solutions, and their equivalents, which may be, but is notlimited to, the above-mentioned intraocular irrigating-washing solutions(e.g., Opeguard MA and Opeguard Neo Kit, both manufactured by SenjuPharmaceutical Co., Ltd.), or an artificial tear or their equivalents.

In the case where the transparent tissue-visualizing preparation of thepresent invention is in the form of a suspension, the content of thefine particles of a macromolecular compound in the suspension, forachieving a steady effect of enhancement of visibility, is preferably inthe range of 0.005-10 w/v %, more preferably 0.01-5 w/v %, andparticularly preferably 0.1-2 w/v %. However, the content may falloutside these ranges, for a content lower than these, such as 0.0001 w/v%, would be capable of providing certain visibility, and a highercontent may also be employed insofar as it causes no inconvenience inhandling, e.g., in dispersing the fine particles. For convenience, thecontent may generally be set in the range of 0.01-5 w/v % as desired.Where one or more of the aforementioned polyvinyl-based compounds orpolyols are added to increase dispersibility of the fine particles, theconcentration of any of them may be set as desired in the range of 0.1-5w/v %, though it may fall outside this range, since even a lower contentwould provide some effect corresponding to the content, and a highercontent would bring no particular disadvantage.

The transparent tissue-visualizing preparation of the present inventionmay be provided, as aforementioned, in the form of a powder or, instead,a suspension consisting of aqueous medium and fine particles dispersedin it. Furthermore, it may be provided in such a form that comprises apowder consisting of fine particles of a macromolecular compound (and,as desired, one or more additives such as polyvinyl-based compounds,polyols and the like) and an aqueous medium in which to disperse thepowder to make a suspension prior to use in a surgical operation, thepowder and the aqueous medium being enclosed, separately and in a mannerof keeping from contacting with each other, in a single mixing anddischarging means. In such a case, an aqueous medium may be one of thoseaforementioned above.

In the present invention, there is no specific limitation as to a“mixing and discharging means” insofar as it allows mixing, bymanipulation from outside, the powder and the aqueous medium separatelyenclosed in it, and allows discharging, by manipulation from outside,thus formed mixture to the outside. As such, there are well known avariety of two-compartment syringes. A two-compartment syringe istypically of a cylindrical body at the front end which is defined adischarge flow path around which an injection needle can be (or alreadyis) fitted, and through the other end of which is received aliquid-tightly inserted piston, and at an intermediate region of whichis placed a liquid-tightly inserted partition that is slidable in thelongitudinal direction, thus forming two chambers, front and rear,within the cylindrical body. Forward of partition is provided, byforming a recess in the interior surface of the cylinder, an elongatedbypassing flow path which extends in the longitudinal direction in theregion greater in length than the thickness of the partition. In thefront chamber is generally enclosed a dried composition, e.g., in theform of a powder, and in the rear chamber a liquid composition (e.g., abuffered solution) as a medium to be mixed with the dried composition.Mixing and discharging is performed as follows. First, the pistoninserted in the rear end is forced to advance and, under a hydraulicpressure created by this, the slidable partition is then advancedforwardly to reach the central region of the bypassing flow path. Thepiston is forced to advance further until the liquid compositioncontained in the rear chamber is expelled into the front chamber throughthe bypassing flow path, and the contents are thus mixed within thefront chamber, and then the mixture is discharged through the dischargeflow path by further advancing the slidable partition through pressingthe piston. It is of particular advantage to provide thetransparent-visualizing preparation of the present invention in a formwhere the preparation is enclosed, separately and in a manner of keepingthe components from contacting with each other, in such a single mixingand discharging means, for such a form greatly enhances convenience inusing the preparation in a surgical operation.

The transparent tissue-visualizing preparation of the present invention,as desired, may contain: pharmaceutically acceptable additives, such asisotonizers (salts like sodium chloride, potassium chloride, etc.;saccharides like glycerol, glucose, etc.; polyols like sorbitol,mannitol, propylene glycol, etc.,; boric acid, borate, etc.), bufferingagents (phosphate buffer, acetate buffer, borate buffer, carbonatebuffer, citrate buffer, tris buffer, etc.), thickening agents(hydroxyethylcellulose, hydroxypropylcellulose,hydroxypropylmethylcellulose, carboxycellulose sodium, polyvinylalcohol,polyvinylpyrrolidone, polyethylene glycol, sodium alginate, etc.),stabilizers (sodium bisulfite, ascorbic acid, sodium ascorbate,dibutylhydroxytoluene, etc.), pH adjusting agents (hydrochloric acid,sodium hydroxide, phosphoric acid, acetic acid, etc.), and the like.Furthermore, insofar as it does not conflict with the purpose of thepresent invention, a pharmacologically active ingredient may also becontained in the transparent tissue-visualizing agent of the presentinvention. The transparent tissue-visualizing agent of the presentinvention, when provided in the form of a suspension (or in the formwhere a powder and an aqueous medium are separately enclosed), its pH(or pH after mixing) is generally adjusted to fall in the range of4.0-8.0, preferably about 5.0-7.5.

The present invention also provides a method for improving visibility ofa transparent tissue of the eye, which method comprises bringing acomposition comprising the fine particles as described above intocontact with a transparent tissue of the eye, as well as use of the fineparticles as described above for the production of a transparenttissue-visualizing preparation which is designed to be brought intocontact with a transparent tissue of the eye to visualize it.

EXAMPLES

The present invention will be described in further detail with referenceto examples, comparison examples and test examples. However, it is notintended that the present invention be limited to those examples.

Test Example 1 Test for Visibility Improvement of the Vitreous Body

Pigmented rabbits were euthanized and the eyeballs (6 from threerabbits) were enucleated. The cornea was excised by giving a cut withscissors along the limbus. The iris and the lens were removed to leavean optic cup behind, which was spread by giving it a crucial incisionfrom the anterior side. Vitreous body that flowed out was removed andthe retina was exposed, with vitreous body attached to part of it. Anaqueous preparation (suspension) containing 1 w/v % of soluble starchwas prepared according to the formula below, about 100 μl of which wasapplied in drops onto that part of the retina. Using an opticalmicroscope, observations were made of the part onto which thepreparation was applied in drops. As a result, fine particles of thesoluble starch were found adhering to the vitreous body in all of the 6eyes, and the visibility of the vitreous body was thereby improved.

(Formula) Soluble starch 1.0 g Opeguard MA to 100 mL(For Opeguard MA, see Test Materials and Reagents below)

Test Example 2 Confirmation of Visibility on Enucleated Pig Eye

1. Tissue used:

Pig eyeballs purchased from a meat processing facility were used.Immediately after purchase, tissues surrounding the pig eyeballs werecut off and removed, and the eyeballs thus isolated were immersed in asolution of low molecular weight dextran (Low Molecular Weight Dextran LInjection, Otsuka Pharmaceutical Co., Ltd.) until the test was started.

2. Test Materials and Reagents:

The following test materials and reagents were used.

(1) Soluble starch (manufactured by Nacalai Tesque, Inc., Reagent Code33131-42, Spec. GR (Guaranteed Reagent))

(2) Chitosan (manufactured by Funakoshi Co., Ltd., Reagent Code K-03,

Particle size: 30 mesh pass—95% or more)

(3) PLA-0005 (manufactured by Wako Pure Chemical Industries, Ltd., D,L-polylactic acid (MW 5000))

(4) PLA-0010 (manufactured by Wako Pure Chemical Industries, Ltd., D,L-polylactic acid (MW 10000))

(5) PLGA-5005 (manufactured by Wako Pure Chemical Industries, Ltd.,lactic acid-glycolic acid copolymer (MW 5000))

(6) Povidone K-30 (manufactured by BASF Japan Ltd., JapanesePharmacopoeia)

(7) Gosenol EG-05 (manufactured by Nippon Synthetic Chemical Industries,Co., Ltd., polyvinylalcohol EG-05)

(8) D-mannitol (manufactured by Nacalai Tesque, Inc.)

(9) Opeguard MA (manufactured by Senju Pharmaceutical Co., Ltd.,intraocular-irrigating solution, containing (per 1 mL): glucose 1.5 mg,sodium chloride 6.6 mg, potassium chloride 0.36 mg, calcium chloride0.18 mg, magnesium sulfate 0.3 mg, sodium bicarbonate 2.1 mg)

3. Method for Preparation of Transparent Tissue-Visualizing Preparation:

3-1. Preparation of transparent tissue-visualizing preparations ofComparison Example 1 and Examples 1-5

See Table 1. PLA 0005 was gradually introduced into an A-O Jet Mill(Seishin Enterprise Co., Ltd.) and pulverized (Pressure conditions:Grinding Nozzle 0.4 MPa, Pusher Nozzle 0.4 MPa). To the powder thusobtained was added Opeguard MA to prepare 2% and 1% suspensions, whichwere made Examples 5 and 4 suspensions, respectively. The Example 4suspension was diluted tenfold with Opeguard MA to make Example 3suspension. The Example 3 suspension was diluted tenfold with OpeguardMA to make Example 2 suspension. The Example 2 suspension was dilutedtenfold with Opeguard MA to make Comparison Example 1 suspension. Ten mLof the 1% suspension of Example 4 was centrifuged (1000 rpm, 5 min) tospin down large particles, and the supernatant was collected to makeExample 1 suspension. One mL of the supernatant was taken and, based onits dry weight, the relative content of PLA 0005 fine particles in thesupernatant was determined (in this calculation, the weight of the solidoriginally contained in Opeguard MA was subtracted).

3-2. Preparation of Transparent Tissue-Visualizing Preparation ofComparison Examples 3-5 and Examples 9-25

See Tables 2-4. To a volume of purified water was added one of thefollowing reagents so that the indicated concentration of it wasattained, and subjected to filtration through 0.22 μm hydrophilic filterto make solution A in the corresponding example: 1.11% of Povidone K-30(, Examples 11-13 and 22-25, upon preparation), 1.11% of D-mannitol(Examples 17 and 22-25, upon preparation), and 0.22% for Gosenol EG-05(Example 25, upon preparation). A solution of 10% PLA0005 was prepared(acetone/ethanol=4/6) and filtered through a 0.22 μm hydrophobic filterto make solution B. Opeguard MA was made as solution C.

The solution A for each Example and the solution B, a poly lactic acidsolution, were mixed at the ratio of 9:1 in the following manner.Briefly, to the solution A, while stirred at 700-800 rpm with a stirrer,was added dropwise the solution B at a rate of about 100 μL/sec to allowPLA0005 to precipitate as fine particles. This was stirred for about 30minutes, and aggregation product was removed through a sieve (mesh size106 μm) to make suspension D. This, through lyophilization, gave apowder form sample E. The powder form sample E was dispersed in thesolution C so that the final content of the fine particle reached 1 w/v% to make compositions of Examples 11, 17 and 22. The composition ofExample 11 containing 1 w/v % of the fine particles was sequentiallydiluted with Opeguard MA, tenfold at a time, until a content of 0.001%was reached, and a composition of which the content is 0.1 w/v % wasmade Example 10, 0.01 w/v % Example 9, and 0.001 w/v % ComparisonExample 3. The composition of Example 17 was sequentially diluted inlike manner with Opeguard MA, tenfold at a time, and a composition ofwhich the content was 0.1 w/v % was made Example 16, 0.01 w/v % Example15, 0.001 w/v % Example 14, and 0.0001 w/v % Comparison Example 4. Thecomposition of Example 22 was sequentially diluted in like manner withOpeguard MA, tenfold at a time, and a composition of which the contentwas 0.1 w/v % was made Example 21, 0.01 w/v % Example 20, 0.001 w/v %Example 19, 0.0001 w/v % Example 18, and 0.00001 w/v % ComparisonExample 5. The powder form sample E, which was used in Example 11, wasdispersed in the solution C so that the final content of PLA0005 reached2 w/v % or 5 w/v % to make suspensions of Examples 12 or 13,respectively. Furthermore, the powder form sample E, which was used inExample 22, was dispersed in the solution C so that the final content ofPLA0005 reached 2 w/v % or 5 w/v % to make suspensions Examples 23 and24, respectively. The powder form sample E which had been preparedthrough precipitation in the Gosenol EG-05-containing aqueous solution,was dispersed in the solution C so that the final content of PLA0005 was1 w/v % to make a suspension of Example 25.

Lyophilization in the process for preparation of the compositions ofExamples 11-13, Example 17 and Examples 22-25 was carried out asfollows. The suspension D was frozen by storing it at −40° C. for 6hours and, after the pressure was reduced to not higher than 100 μmHg at−40° C., dried for at least 24 hours. The temperature then was allowedto elevate from −40° C. at a rate of 10° C./hour until it reached +20°C. Further drying at +20° C. at not higher than 100 μmHg for 24 hoursgave the powder sample.

3-3. Preparation of Transparent Tissue-Visualizing Preparations ofComparison Example 2 and Examples 6-8

Ten mL of the suspension of Example 11 prepared in the same manner asdescribed in 3-2 above was centrifuged (conditions: 1000 rpm, 5 min) tospin down large particles and supernatant was collected. The supernatantwas made a composition of Example 8. The composition of Example 8 wastenfold diluted with Opeguard MA to make a composition of Example 7,which in turn was tenfold diluted with Opeguard MA to make a compositionof Example 6, which then was tenfold diluted with Opeguard MA to make acomposition of Comparison Example 2. The content of PLA0005 in thesupernatant was determined based on the dry weight of 1 mL of thesupernatant of the composition of Example 8. In this calculation, theweight of Povidone and the solid contained in Opeguard MA wassubtracted.

3-4. Preparation of the Transparent Tissue-Visualizing Preparations ofComparison Example 6 and Examples 26-31

Referring to Table 5, soluble starch was gradually put into the A-O JetMill (Seishin Enterprise Co., Ltd.) and pulverized (Pressure conditions:Grinding Nozzle 0.4 MPa, Pusher Nozzle 0.4 MPa). To this sample wasadded Opeguard MA to prepare a suspension whose content of the fineparticles was 10 w/v %, which was made a suspension of Example 31. Thesuspension of Example 31 was diluted with Opeguard MA, and a 1 w/v %suspension thus prepared was made Example 30, a 0.1 w/v % suspensionExample 29, a 0.01 w/v % suspension Example 28, a 0.005 w/v % suspensionExample 27, a 0.0025 w/v % suspension Example 26, and a 0.001 w/v %suspension Comparison Example 6.

3-5. Preparation of Transparent Tissue-Visualizing Preparations ofExample 32-35:

Referring to Table 6, PLGA5005 or chitosan was gradually put into theA-O Jet Mill (Seishin Enterprise Co., Ltd.) and pulverized (Pressureconditions: Grinding Nozzle 0.4 MPa, Pusher Nozzle 0.4 MPa). To eachthese was added Opeguard MA to prepare 1 w/v % suspensions, and thePLGA5005 containing suspension was made Example 33, and thechitosan-containing suspension was made Example 35. And a 1 w/v %PLA0010 suspension was prepared by addition of Opeguard and made Example34. The suspension of Example 33 was centrifuged (conditions: 1000 rpm,5 min) to spin down large particles and supernatant was collected. Thesupernatant was made Example 32. The content of PLGA5005 in thesupernatant was determined based on the dry weight of 1 mL of thesupernatant. In this calculation, the weight of the solid contained inOpeguard MA was subtracted.

4. Test Method:

The average size of the fine particles in Example 1, Example 4, Example8, Example 11, Example 17, Example 22, Example 30 and Examples 32-35 wasmeasured using a laser diffraction particle size analyzer (SALD-2100,Shimadzu Corporation). The average size of the fine particles wasdefined as the average value that was calculated by the method shown inNote 1. Briefly, over the full range of the particle size distribution,the size of the particles falling within each of the intervals (x_(j),x_(j+1)) created by dividing the range by a large number (n) wasrepresented by the mean (in logarithm) of the logarithms of the particlesizes at the both end of the interval, and each of them were multipliedby the % difference in volume of all the particles detected in the veryinterval relative to the volume of those found in the entire range ofdistribution (i.e., frequency in the volume distribution), and thevalues thus obtained with regard to all the intervals, which togethercovered the entire range of distribution, were added and the average wastaken, and, from the average (μ), the average value of the size (10μ) ofthe fine particles was calculated. In the case of Examples wheredispersion of fine particles by shaking was found difficult due to theiraggregation, measurement was made after they were ultrasonicallydispersed. For other Comparison Examples and Examples than thosedescribe above, no measurement of the particle size was made, since thepowders used in them were of the same lots as those employed in theabove described samples.(Note 1) Method for Calculation of the Average Particle Size:$\mu = {\frac{1}{100}{\sum\limits_{j = 1}^{n}{q_{j}\frac{{\log_{10}x_{j}} + {\log_{10}x_{j - 1}}}{2}}}}$[MATH 1]where x_(j): particle size (μm)

-   -   q_(j): % difference in volume (frequency in the volume        distribution)

For performing vitreous surgery, a surgical instrument for vitreoussurgery (Ocutome (registered trademark), ALCON) and a microscope forophthalmic surgery (CARL ZEISS) were used. A pig eyeball was incised ata point outside the limbus, about 3 mm away from it, with a knife forophthalmic surgery (20G V-Lance™, ALCON), and the sclera was cutparallel to the limbus to create a port for an irrigation cannula. Anirrigation cannula was inserted through the port thus created, andphysiological saline (Otsuka Pharmaceutical Co., Ltd.) was constantlyinfused during the surgical operation. With a knife for ophthalmicsurgery (20G V-Lance™, ALCON), an incision parallel to the limbus wasmade to the sclera at a point about 3 mm outside of the limbus and atabout 120°, around the center of the cornea, away from the intraocularirrigation port, to create a port for vitreous cutter. A vitreous cutterwas inserted through the port thus created, and the lens and othertissues surrounding it were removed and, after it was confirmed througha microscope that the ocular fundus was visible, a central portion ofthe vitreous body was removed. Using a disposable syringe (5 mL TerumoCorporation) equipped with a non bevel needle (22G, Terumo Corporation),1 mL of a transparent tissue-visualizing preparation was infused,through the port for a vitreous cutter, into the cavity which had beencreated by removal of the central portion of the vitreous body. Afterthe fine particles drifting in the liquid was removed by suctioning withthe vitreous cutter, a light guide was inserted through the port for avitreous cutter and the visibility of the vitreous body due to theadhered fine particles were assessed by examining the anterior part(lenticular side), equatorial part and the posterior part of thevitreous body (retinal fundus side), respectively, according to thefollowing criterion. Transparent tissue-visualizing preparations wereassessed for their visualizing ability/inability, as well as the degreeof it, and judged to possess a practical level of visualizing effect ifscore 2 or 3 of the assessment criterion table was obtained with atleast one of these parts.

<Assessment Criterion>

0 - - - No fine particle was observed adhering to the remaining vitreousbody, and the shape of the vitreous body is not visible, either.

1 - - - Fine particles were observed adhering to the remaining vitreousbody, but the shape of the vitreous body is not visible.

2 - - - The shape of the remaining vitreous body is visible due toadhering fine particles, with the retina clearly visible through theadhering fine particles.

3 - - - The shape of the remaining vitreous body is visible due toadhering fine particles, with the retina only dimly visible through theadhering particles.

5. Test Results:

5-1. Visualizing Ability of PLA-0005-Containing TransparentTissue-Visualizing Preparations:

The results of the test on visibility improving effect ofPLA-0005-containing transparent tissue-visualizing preparations areshown in Tables 1-4. To prepare PLA0005 of different particle sizes,PLA0005 was pulverized using the A-O Jet Mill for Comparison Example 1and Examples 1-5. For Comparison Examples 2-5 and Examples 6-25, fineparticles were prepared by dissolving PLA0005 in an organic solvent andthen causing precipitation to take place as fine particles. PLA0005 wasprecipitated as fine particles in a Povidone solution for ComparisonExamples 2 and 3 and Examples 6-13, in a mannitol solution forComparative Example 4 and Examples 14-17, in a solution of Povidone andmannitol for Comparative Example 5 and Examples 18-24, and in a solutionof Gosenol and mannitol for Example 25. For Comparative Example 2 andExample 1 and Examples 6-8, a high content PLA0005 suspension wascentrifuged and supernatant was collected to obtain finer particles. Asa result of these processes, fine particles were obtained whose averageparticle size fell in the range of 0.8 μm-60.3 μm. For these fineparticles, concentrations giving useful vitreous body-visualizingpreparations were determined in the test of a visualization effect, and,as a result, it was found that the fine particles that exhibited avisualization effect at the lowest content was those of PLA0005 of anaverage particle size of 17.9 μm, which was useful down to the contentas low as 0.0001 w/v % (Example 18). PLA0005 particles of the smallestparticle size of 0.8 μm of the samples were found useful down to thecontent as low as 0.0073 w/v % (Example 6). PLA0005 particles of thelargest size of 60.3 μm were found useful down to the content as low as0.001% (Example 14). Particles of the sizes of 17.9 and 22 μm were foundto be dispersed even at a content of as high as 5%, with goodvisualizing effect, thus were found useful as transparenttissue-visualizing preparations (Example 13 and Example 24). When thecontent of PLA0005 was 2%, particles of any of the sizes tested werefound usable as transparent tissue-visualizing preparations (Example 5,Example 12 and Example 23). From the findings as noted above, it isevident that the tested polylactic acid fine particles could exhibit avisibility improving effect even at a content of 0.0001 w/v %, and willunfailingly exhibit the effect at a content of not less than 0.005 w/v%. TABLE 1 Results of test on visibility improving effect ofPLA-0005-containing transparent tissue-visualizing preparationsComparison Comparison Example 1 Example 1 Example 2 Example 3 Example 4Example 5 Example 2 Example 6 PLA-0005 (g) 0.36 0.001 0.01 0.1 1 20.00073 0.0073 Povidone (g) — — — — — — 0.00073 0.0073 D-Mannitol (g) —— — — — — — — Opeguard MA q.s. q.s. q.s. q.s. q.s. q.s. q.s. q.s. (to100 mL) Average 1.9 6.2 6.2 6.2 6.2 6.2 0.8 0.8 particle size (μm)Visibility Anterior 2 1 2 2 3 3 1 2 part Posterior 1 1 1 2 3 3 1 2 partEquatorial 2 1 2 2 3 3 1 2 part Sum 5 3 5 6 9 9 3 6

TABLE 2 Results of test on visibility improving effect ofPLA-0005-containing transparent tissue-visualizing preparationsComparison Example 7 Example 8 Example 3 Example 9 Example 10 Example 11Example 12 Example 13 PLA-0005 (g) 0.073 0.73 0.001 0.01 0.1 1 2 5Povidone (g) 0.073 0.73 0.001 0.01 0.1 1 2 5 D-Mannitol (g) — — — — — —— — Opeguard MA q.s. q.s. q.s. q.s. q.s. q.s. q.s. q.s. (to 100 mL)Average 0.8 0.8 22 22 22 22 22 22 particle size (μm) Visibility Anterior2 3 1 2 3 3 3 3 part Posterior 2 3 1 2 3 3 3 3 part Equatorial 2 3 1 2 33 3 3 part Sum 6 9 3 6 9 9 9 9

TABLE 3 Results of test on visibility improving effect ofPLA-0005-containing transparent tissue-visualizing preparationsComparison Comparison Example 4 Example 14 Example 15 Example 16 Example17 Example 5 Example 18 Example 19 PLA-0005 (g) 0.0001 0.001 0.01 0.1 10.00001 0.0001 0.001 Povidone (g) — — — — — 0.00001 0.0001 0.001D-Mannitol (g) 0.0001 0.001 0.01 0.1 1 0.00001 0.0001 0.001 Opeguard MAq.s. q.s. q.s. q.s. q.s. q.s. q.s. q.s. (to 100 mL) Average particle60.3 60.3 60.3 60.3 60.3 17.9 17.9 17.9 size (μm) Visibility Anterior 12 2 2 3 1 2 2 part Posterior 0 1 2 2 3 0 1 1 part Equatorial 1 2 2 2 3 12 2 part Sum 2 5 6 6 9 2 5 5

TABLE 4 Results of test on visibility improving effect of PLA-0005-containing transparent tissue-visualizing preparations Example 20Example 21 Example 22 Example 23 Example 24 Example 25 PLA-0005 (g) 0.010.1 1 2 5 1 Povidone (g) 0.01 0.1 1 2 5 — Gosenol (g) — — — — — 0.2D-Mannitol (g) 0.01 0.1 1 2 5 1 Opeguard MA q.s. q.s. q.s. q.s. q.s.q.s. (to 100 mL) Average particle 17.9 17.9 17.9 17.9 17.9 11.9 size(μm) Visibility Anterior 2 2 3 3 3 3 part Posterior 1 2 3 3 3 3 partEquatorial 2 2 3 3 3 3 part Sum 5 6 9 9 9 95-2. Results of Test on Visibility Improving Effect of SolubleStarch-Containing Transparent Tissue-Visualizing Preparations

Referring to Table 5, soluble starch was pulverized using A-O Jet Millto obtain particles of the size of 35.8 μm. The 35.8 μm particlessuccessfully visualized vitreous body at the content down to 0.0025 w/v% (Example 26). Therefore, they can be used as a transparenttissue-visualizing preparation at a content falling in the range of atleast from 0.0025% to 10 w/v %. TABLE 5 Results of test on visibilityimproving effect of soluble starch-containing transparenttissue-visualizing preparations Comparison Example 6 Example 26 Example27 Example 28 Example 29 Example 30 Example 31 Soluble starch (g) 0.0010.0025 0.005 0.01 0.1 1 10 Opeguard MA q.s. q.s. q.s. q.s. q.s. q.s.q.s. (to 100 mL) Average particle 35.8 35.8 35.8 35.8 35.8 35.8 35.8size (μm) Visibility Anterior 1 2 2 1 2 2 3 part Posterior 1 1 1 2 2 3 3part Equatorial 1 1 2 1 2 2 3 part Sum 3 4 5 4 6 7 95-3. Results of Test on Visibility Improving Effect of TransparentTissue-Visualizing Preparations Containing Other MacromolecularCompounds

Referring to Table 6, vitreous body was made visible with the use of thefine particles consisting of PLGA5005, PLA0010 or chitosan, at a contentof 1 w/v % (Examples 33-35). PLGA5005 at a content of 0.01% could alsovisualize the vitreous body (Example 32). TABLE 6 Results of test onvisibility improving effect of transparent tissue- visualizingpreparations containing other macromolecular compounds Example ExampleExample Example 32 33 34 35 PLGA-5005 (g) 0.01 1 — — PLA-0010 (g) — — 1— Chitosan (g) — — — 1 Opeguard MA q.s. q.s. q.s. q.s. (to 100 mL)Average particle size 3.1 15.5 65.4 133.9 (μm) Visibility Anterior 2 3 23 part Posterior 2 3 2 3 part Equatorial 2 3 2 3 part Sum 6 9 6 9

Test Example 2 Test of Redispersibility of TransparentTissue-Visualizing Preparations

1. Test Method:

See Table 7. Transparent tissue-visualizing preparations of Examples36-38 were prepared in the same manner as in Test Example 1, In a 5-mLsyringe, 0.02 g of the powder sample prepared in Example 36, 0.04 g ofthe powder sample prepared in Example 37 or 0.06 g of the powder sampleprepared in Example 38 was mixed with 2 mL of the following dispersionmedium and shaken by hand, and the number of the shakes that wasrequired for the powder to be dispersed in the dispersion medium wascounted. <Formula of dispersion medium> Sodium chloride 0.75 g Potassiumchloride 0.16 g Sodium hydrogen phosphate  0.1 g Sodium dihydrogenphosphate 0.015 g  Hydrochloric acid q.s. (pH 7) Purified water to 100mL

TABLE 7 Redispersibility of preparations containing Povidone andD-mannitol Example 36 Example 37 Example 38 PLA-0005 (g) 1 1 1D-Mannitol (g) — 1 1 Povidone (g) — — 1 Dispersion q.s. q.s. q.s.medium* (to 100 mL) Number of 200 or 15  7 shakes needed more until thepowder was dispersed*A phosphate buffer (pH 7) was employed containing 0.75% of sodiumchloride, and 0.16% of potassium chloride.2. Test Results:

The number of shakes required to achieve dispersion of the PLA0005 fineparticles after their mixing with the medium was reduced by addition ofmannitol, indicating that redispersibility was improved. When Povidonewas further added to the mixture, the number of shakes required toachieve dispersion was further reduced, showing that further improvementin redispersibility was obtained.

Production Example 1 Prefilled Vial-Type Transparent Tissue-VisualizingPreparation

An aqueous solution containing 1.11 w/v % of D-mannitol and 1.11 w/v %of Povidone K-30 was filtered through a hydrophilic filter having thepore size of 0.22 μm to make a solution A. An acetone solution of 10 w/v% of PLA0005 (acetone/ethanol=4/6) was prepared and filtered through ahydrophobic filter having the pore size of 0.22 μm to make a solution B.Opeguard MA was made a solution C.

The solutions A and B were mixed at a proportion of 9:1 in the followingmanner. Briefly, to the solution A, stirred at 700-800 rpm with astirrer, was added the solution B, at a rate of about 100 μL/sec, to letPLA0005 precipitate as fine particles. The mixture was stirred for about30 minutes, and aggregation products were removed through a sieve (meshsize 106 μm) to obtain a suspension D. Lyophilization of this in a vialgave a powder form sample E. Prior to use, the solution C was pouredinto the vial containing the powder form sample E to prepare a sample.

Lyophilization was carried out in the following manner. Briefly, thesuspension D was frozen by storing it at −40° C. for 6 hours and then,after the pressure was reduced to 100 μmHg at −40° C., dried for atleast 24 hours. The temperature then was allowed to elevate from −40°C., at a rate of 10° C./hour, until it reached +20° C. The dryingprocess was further continued at +20° C. and below 100 μmHg for at least24 hours.

Production Example 2 Prefilled Double Chamber Syringe-Type TransparentTissue-Visualizing Preparation

An aqueous solution containing 1.11 w/v % of D-mannitol and 1.11 w/v %of Povidone K-30 was filtered through a hydrophilic filter having thepore size of 0.22-μm to make a solution F. On the other hand, a 10 w/v %solution of PLA0005 (acetone/ethanol=4/6) was filtered through ahydrophobic filter having the pore size of 0.22 μm to make a solution G.A phosphate buffer (pH 7) containing 0.75% of sodium chloride and 0.16%of potassium chloride was made a liquid H.

The solutions F and G were mixed at a proportion of 9:1 in the followingmanner. Briefly, to the solution F, stirred at 700-800 rpm with astirrer, was added the solution G, at a rate of about 100 μL/sec, toallow PLA0005 to precipitate as fine particles. The mixture was stirredfor about 40 minutes (for 10 minutes of which a reduced pressure wasapplied), and aggregation products were removed through a sieve (meshsize 106 μm) to obtain a suspension J. Two ml each of this was dispensedon the solid phase side 3 in the double chamber syringe 1, which isschematically illustrated in FIG. 1, and subjected to lyophilization togive powder L. By fitting a rubber plug, which served as the slidablepartition 2 between the solid phase side 3 and the liquid phase side 4,filling the syringe, on the liquid phase side 4, with 2 mL of the liquidH serving as the medium 6, and then fitting a plug serving as the piston8, a prefilled double chamber syringe-type transparenttissue-visualizing preparation was obtained. The numeral 9 indicates alongitudinal bypassing flow path, which is defined by a partial recessin the interior wall of the double chamber syringe 1. Lyophilization wascarried out in the following manner. Briefly, the suspension J wasfrozen by storing it at −40° C. for 6 hours and, after the pressure wasreduced to 100 μmHg at −40° C., dried for at least 24 hours. Thetemperature then was allowed to elevate from −40° C., at a rate of 10°C./hour, until it reached +20° C. The drying process was furthercontinued at +10° C. and below 100 μmHg for at least 24 hours.

The transparent tissue-visualizing preparation of this example is usedin the following manner. Briefly, the piston 8 is pushed in to advance,and, utilizing the pressure generated by this in the medium liquid Henclosed on the liquid phase side 4, the slidable partition 2 then ispushed to advance. When the rear edge of the slidable partition 2reaches the bypassing flow path 9, the liquid phase side 4 and the solidphase side 3 are placed into communication with each other, and theliquid H starts to flow into the solid side 3. By further pushing in thepiston 8 until it abuts on the slidable partition 2, all the liquid H istransferred to the solid phase side 3, where it mixes with the powder L.After completion of the mixing, the piston 8 is further pushed in(together with the sidable partition 2) to advance, and the mixtureliquid then is discharged from the discharge flow path 7 (through aneedle, etc. not shown) to the part on which a surgical operation isbeing performed.

Preparation Example 1 Transparent Tissue-Visualizing PreparationContaining Soluble Starch Fine Particles

A transparent tissue-visualizing preparation of the following formulawas prepared in a conventional manner. In the preparation, the averageparticle size of the soluble starch fine particles was about 50 μm.Solubile starch 1.0 g Opeguard MA to 100 mL

Preparation Example 2 Transparent Tissue-Visualizing PreparationContaining Chitosan Fine Particles

A transparent tissue-visualizing preparation of the following formulawas prepared in a conventional manner. Chitosan 1.0 g Opeguard MA to 100mL

Preparation Example 3 Transparent Tissue-Visualizing PreparationContaining Polylactic Acid Fine Particles

A transparent tissue-visualizing preparation of the following formulawas prepared in a conventional manner. In the preparation, the averageparticle size of the polylactic acid fine particles was about 0.2-0.3μm. Polylactic acid 1.0 g Opeguard MA to 100 mL

Preparation Example 4 Transparent Tissue-Visualizing PreparationContaining Lactic Acid-Glycolic Acid Copolymer Fine Particles

A transparent tissue-visualizing preparation of the following formulawas prepared in a conventional manner. In the preparation, the averageparticle size of the lactic acid-glycolic acid copolymer fine particleswas about 0.06-0.07 μm. Lactic acid-glycolic acid copolymer 1.0 gOpeguard MA to 100 mL

Preparation Example 5 Transparent Tissue-Visualizing PreparationContaining Soluble Starch Fine Particles

A transparent tissue-visualizing preparation of the following formulawas prepared in a conventional manner. Soluble starch 1.0 g Sodiumdihydrogen phosphate, dihydrate 0.10 g  Sodium chloride 0.9 g Sodiumhydroxide q.s. Purified water to 100 mL pH 7.0

Preparation Example 6 Transparent Tissue-Visualizing PreparationContaining Chitosan Fine Particles

A transparent tissue-visualizing preparation of the following formulawas prepared in a conventional manner. Chitosan 0.10 g Sodium chloride0.55 g Potassium chloride 0.16 g Exsiccated sodium carbonate 0.06 gSodium hydrogen phosphate 0.18 g Boric acid  1.2 g Borax q.s. Sterilepurified water to 100 mL pH 7.3

Preparation Example 7 Transparent Tissue-Visualizing PreparationContaining Polylactic Acid Fine Particles

A transparent tissue-visualizing preparation of the following formulawas prepared in a conventional manner. Polylactic acid  10 g Sodiumdihydrogen phosphate, dihydrate 0.1 g Sodium chloride 0.9 g Sodiumhydroxide q.s. Sodium edetate 0.1 g Sterile purified water to 100 mL pH7.0

Preparation Example 8 Transparent Tissue-Visualizing PreparationContaining Lactic Acid-Glycolic Acid Copolymer Fine Particles

A transparent tissue-visualizing preparation of the following formulawas prepared in a conventional manner. Lactic acid-glycolic acidcopolymer 0.10 g Sodium chloride 0.55 g Potassium chloride 0.16 gExsiccated sodium carbonate 0.06 g Sodium hydrogen phosphate 0.18 gBoric acid 1.2 g Borax q.s. Hydroxypropylmethylcellulose 0.1 g Sterilepurified water to 100 mL pH 7.3

INDUSTRIAL APPLICABILITY

In surgical operations on transparent tissues of the eye, i.e., thevitreous body, the lens and the cornea, the present invention as definedabove greatly enhances visibility of transparent tissues, whichotherwise are hardly visible in the operative field, neither causingunnecessary pharmacological reactions nor side effects in the body,thereby allowing easier manipulation in such surgical operations and,therefore, making it easier to unfailingly achieve the purpose of thesuch surgical operations.

1. A transparent tissue-visualizing preparation designed to be broughtinto contact with transparent tissues of the eye to enhance visibilitythereof, comprising fine particles of a macromolecular compound.
 2. Thetransparent tissue-visualizing preparation of claim 1, wherein the fineparticles are those one g of which cannot completely be dissolved inless than 30 mL of water within 30 minutes at 20° C.
 3. The transparenttissue-visualizing preparation of claim 1, wherein the average size ofthe particles is 0.01-500 μm.
 4. The transparent tissue-visualizingpreparation of claim 1, wherein the macromolecular compound is abiodegradable macromolecular compound and/or a water solublemacromolecular compound.
 5. The transparent tissue-visualizingpreparation of claim 4, wherein the biodegradable macromolecularcompound is selected from fatty acid polyesters, polysaccharides andderivatives thereof.
 6. The transparent tissue-visualizing preparationof claim 4, wherein the water soluble macromolecular compound isselected from acrylic polymer compounds and styrene-based polymercompounds.
 7. The transparent tissue-visualizing preparation of claim 1,wherein the macromolecular compound is selected from the groupconsisting of starch, soluble starch, pregelatinized starch, cellulose,acetylcellulose, hydroxypropylmethylcellulose, chitosan, chitin,dextran, polylactic acid, polyglycolic acid, lactic acid-glycolic acidcopolymer, polyhydroxybutyric acid, polyhydroxyvaleric acid,polycaprolactone, hydroxybutyric acid-glycolic acid copolymer, lacticacid-caprolactone copolymer, polyethylene succinate, and polybutylenesuccinate, sodium polyacrylate, sodium methacrylate and sodiumpolystyrenesulfonate.
 8. The transparent tissue-visualizing preparationof claim 7, wherein the average molecular weight of the macromolecularcompound is 500-200000.
 9. The transparent tissue-visualizingpreparation of claim 8 further comprising a polyvinyl-based compoundand/or a polyol.
 10. The transparent tissue-visualizing preparation ofclaim 9 containing, per one part by weight of the fine particles,0.05-10 part by weight of the polyvinyl-based compound and/or 0.05-10part by weight of the polyol.
 11. The transparent tissue-visualizingpreparation of claim 9, wherein the polyvinyl-based compound ispolyvinyl pyrrolidone and/or polyvinylalcohol.
 12. The transparenttissue-visualizing preparation of claim 9, wherein the polyol ismannitol.
 13. The transparent tissue-visualizing preparation of claim 1further comprising an aqueous medium, in which the fine particles aredispersed.
 14. The transparent tissue-visualizing preparation of claim13, wherein the content of the fine particles in the transparenttissue-visualizing preparation is 0.005-10 w/v %.
 15. The transparenttissue-visualizing preparation of claim 13 further containing apolyvinyl-based compound and/or a polyol.
 16. The transparenttissue-visualizing preparation of claim 15 containing thepolyvinyl-based compound at a concentration of 0.1-5 w/v % and/or thepolyol at a concentration of 0.1-5 w/v %.
 17. The transparenttissue-visualizing preparation of claim 1 which is a preparationdesigned for injection.
 18. A transparent tissue-visualizing preparationcomprising a single mixing and discharging means which encloses,separately and in a manner of keeping from contacting with each other, apowder composition comprising the fine particles of claim 1 and anaqueous medium.
 19. A method for improving visibility of a transparenttissue of the eye, which method comprises bringing a compositioncomprising the fine particles of claim 1 into contact with a transparenttissue of the eye.
 20. Use of the fine particles of claim 2 for theproduction of a transparent tissue-visualizing preparation which isdesigned to be brought into contact with transparent tissue of the eyeto visualize it.